CN114644695B - Protein for regulating drought resistance of corn and application thereof - Google Patents

Abstract

The application discloses a protein for regulating drought resistance of corn and application thereof. The application discloses a protein for regulating drought resistance of corn, which is named ZmYAB15 and has an amino acid sequence of SEQ ID No. 3. After the ZmYAB15 coding gene is overexpressed in corn, transgenic plants are easier to wilt under drought treatment conditions than control plants, and the survival rate is reduced after rehydration, so that the gene can reduce the drought resistance of plants, and the gene can be used for cultivating plants with reduced drought resistance so as to screen substances capable of improving the drought resistance of the plants.

Description

Protein for regulating drought resistance of corn and application thereof

Technical Field

The application relates to a protein for regulating drought resistance of corn and application thereof in the field of biotechnology.

Background

For a long time, successor scientific researchers have focused on improving stress resistance of plants by various biological means, wherein the traditional breeding mode needs to identify and combine excellent characters, accuracy and efficiency determine success and failure of breeding, and the breeding cycle is long despite high biological safety and stability, and the result is unpredictable. In recent years, molecular breeding from wind-induced water has obvious advantages, mainly including high efficiency, strong pertinence, short period and strong predictability. The improvement of crops by means of genetic engineering provides a new opportunity for rapidly solving the food shortage caused by environmental stress.

Corn is one of three world food crops, the yield of the corn directly affects each corresponding industrial chain, and the adverse stress has a great influence on the yield of the corn, so that the mining of drought resistance genes as much as possible has great strategic significance for improving the drought resistance of the corn and weakening the influence of adverse environment on the yield in the future through a genetic engineering means. At present, the sequencing of the B73 corn genome is finished, the genetic transformation technology and the gene editing technology of corn also tend to be mature, and important guarantee is provided for genetic improvement through a genetic engineering means.

Plants, due to their sessile nature, need to function through intracellular, elaborate signal transduction systems when subjected to stress, with the important plant hormones abscisic acid (ABA) and calcium ions playing a vital role. Under drought stress, a series of reactions are initiated in plants, including changes in stomatal movement, regulation of growth and development, etc., to make the plants effective in a diverse environment. In complex signal regulation networks, transcription factors play a very important role, and plants regulate the expression of drought response genes through a series of key transcription factors, so that plants survive in stress.

Disclosure of Invention

The application aims to solve the technical problem of how to improve the drought resistance of corn.

To solve the above technical problems, the present application provides, first, any one of the following applications of proteins or substances regulating the activity or content of the proteins:

d1 Regulating drought resistance of the plant;

d2 Preparing a product for regulating and controlling drought resistance of plants;

d3 Plant breeding;

the protein (which is named ZmYAB 15) is A1), A2) or A3) as follows:

a1 A protein having an amino acid sequence of SEQ ID No. 3;

a2 A protein which is obtained by substituting and/or deleting and/or adding one or more amino acid residues for the amino acid sequence shown in SEQ ID No.3 in the sequence table and has the same function;

a3 A fusion protein obtained by ligating a tag to the N-terminal or/and the C-terminal of A1) or A2).

In order to facilitate purification of the protein of A1), a tag as shown in the following table may be attached to the amino-terminus or the carboxyl-terminus of the protein consisting of the amino acid sequence shown in SEQ ID No.3 of the sequence Listing.

Table: tag sequence

Label (Label)	Residues	Sequence(s)
			Poly-Arg	5-6 (usually 5)	RRRRR
Poly-His	2-10 (usually 6)	HHHHHH
			FLAG	8	DYKDDDDK
Strep-tag II	8	WSHPQFEK
			c-myc	10	EQKLISEEDL

The protein in A2) has 75% or more identity with the amino acid sequence of the protein shown in SEQ ID No.3 and has the same function. The identity of 75% or more is 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98% or 99% identity.

The protein in A2) can be synthesized artificially or can be obtained by synthesizing the coding gene and then biologically expressing.

The coding gene of the protein in A2) above can be obtained by deleting one or several amino acid residues in the DNA sequence shown in SEQ ID No.2 and/or making one or several base pair missense mutations and/or ligating the coding sequences of the tags shown in the above table at the 5 'and/or 3' ends thereof. Wherein the DNA molecule shown in SEQ ID No.2 encodes the protein shown in SEQ ID No. 3.

The application also provides any one of the following applications of the biological material related to ZmYAB 15:

d1 Regulating drought resistance of the plant;

d2 Preparing a product for regulating and controlling drought resistance of plants;

d3 Plant breeding;

the biomaterial is any one of the following B1) to B7):

b1 A nucleic acid molecule encoding ZmYAB 15;

b2 An expression cassette comprising the nucleic acid molecule of B1);

b3 A recombinant vector comprising the nucleic acid molecule of B1) or a recombinant vector comprising the expression cassette of B2);

b4 A recombinant microorganism comprising the nucleic acid molecule of B1), or a recombinant microorganism comprising the expression cassette of B2), or a recombinant microorganism comprising the recombinant vector of B3);

b5 A transgenic plant cell line comprising the nucleic acid molecule of B1) or a transgenic plant cell line comprising the expression cassette of B2);

b6 A transgenic plant tissue comprising the nucleic acid molecule of B1) or a transgenic plant tissue comprising the expression cassette of B2);

b7 A transgenic plant organ comprising the nucleic acid molecule of B1) or a transgenic plant organ comprising the expression cassette of B2).

In the above applications, the nucleic acid molecule of B1) may be B11) or B12) or B13) or B14) or B15) as follows:

b11 A cDNA molecule or a DNA molecule of SEQ ID No.2 in the sequence table;

b12 A cDNA molecule or a DNA molecule shown in SEQ ID No.2 of the sequence Listing;

b13 A DNA molecule shown in SEQ ID No.1 of the sequence table;

b14 A cDNA molecule or genomic DNA molecule having 75% or more identity to the nucleotide sequence defined in b 11) or b 12) or b 13) and encoding ZmYAB 15;

b15 Under stringent conditions with a nucleotide sequence defined by b 11) or b 12) or b 13) or b 14), and a cDNA molecule or genomic DNA molecule encoding ZmYAB15.

Wherein the nucleic acid molecule may be DNA, such as cDNA, genomic DNA, or recombinant DNA; the nucleic acid molecule may also be RNA, such as mRNA or hnRNA, etc.

The nucleotide sequence of ZmYAB15 of the present application can be easily mutated by one of ordinary skill in the art using known methods, such as directed evolution and point mutation. Those artificially modified nucleotides having 75% or more identity with the nucleotide sequence of ZmYAB15 of the present application are derived from the nucleotide sequence of the present application and are equivalent to the sequence of the present application as long as they encode ZmYAB15 and have a protein function.

The term "identity" as used herein refers to sequence similarity to a native nucleic acid sequence. "identity" includes a nucleotide sequence having 75% or more, or 85% or more, or 90% or more, or 95% or more identity with the nucleotide sequence of the protein consisting of the amino acid sequence shown in SEQ ID No.3 of the present application. Identity can be assessed visually or by computer software. Using computer software, the identity between two or more sequences can be expressed in percent (%), which can be used to evaluate the identity between related sequences.

In the above application, the stringent conditions may be as follows: 50℃in 7% Sodium Dodecyl Sulfate (SDS), 0.5M NaPO ₄ Hybridization with 1mM EDTA, rinsing in2 XSSC, 0.1% SDS at 50 ℃; the method can also be as follows: 50℃in 7% SDS, 0.5M NaPO ₄ Hybridization with 1mM EDTA, rinsing in 1 XSSC, 0.1% SDS at 50 ℃; the method can also be as follows: 50℃in 7% SDS, 0.5M NaPO ₄ Hybridization with 1mM EDTA, rinsing in 0.5 XSSC, 0.1% SDS at 50 ℃; the method can also be as follows: 50℃in 7% SDS, 0.5M NaPO ₄ Hybridization with 1mM EDTA, at 50℃and 0.1×SSC, rinsing in 0.1% sds; the method can also be as follows: 50℃in 7% SDS, 0.5M NaPO ₄ Hybridization with 1mM EDTA, rinsing in 0.1 XSSC, 0.1% SDS at 65 ℃; the method can also be as follows: hybridization was performed in a solution of 6 XSSC, 0.5% SDS at 65℃and then washed once with 2 XSSC, 0.1% SDS and 1 XSSC, 0.1% SDS; the method can also be as follows: hybridization and washing the membrane 2 times at 68℃in a solution of 2 XSSC, 0.1% SDS for 5min each time, and hybridization and washing the membrane 2 times at 68℃in a solution of 0.5 XSSC, 0.1% SDS for 15min each time; the method can also be as follows: hybridization and washing of membranes were performed at 65℃in 0.1 XSSPE (or 0.1 XSSC), 0.1% SDS solution.

The 75% or more identity may be 80%, 85%, 90% or 95% or more identity.

In the above applications, the expression cassette described in B2) containing a nucleic acid molecule encoding ZmYAB15 refers to DNA capable of expressing ZmYAB15 in a host cell, and the DNA may include not only a promoter for promoting the transcription of its gene but also a terminator for terminating the transcription of its gene. Further, the expression cassette may also include an enhancer sequence. Promoters useful in the present application include, but are not limited to: constitutive promoters, tissue, organ and development specific promoters, and inducible promoters. Examples of promoters include, but are not limited to: a constitutive promoter of cauliflower mosaic virus 35S; wound-inducible promoters from tomato, leucine aminopeptidase ("LAP", chao et al (1999) Plant Physiol 120:979-992); a chemically inducible promoter from tobacco, pathogenesis-related 1 (PR 1) (induced by salicylic acid and BTH (benzothiadiazole-7-carbothioic acid S-methyl ester); tomato protease inhibitor II promoter (PIN 2) or LAP promoter (both inducible with methyl jasmonate); heat shock promoters (U.S. Pat. No. 5,187,267); tetracycline-inducible promoters (U.S. Pat. No. 5, 057,422); seed-specific promoters, such as the millet seed-specific promoter pF128 (CN 101063139B (China patent 200710099169.7)), seed storage protein-specific promoters (e.g., promoters of phaseolin, napin, oleosin, and soybean beta-cone (Beachy et al (1985) EMBO J. 4:3047-3053)). They may be used alone or in combination with other plant promoters. Reference herein toAll references to (c) are incorporated herein by reference in their entirety. Suitable transcription terminators include, but are not limited to: agrobacterium nopaline synthase terminator (NOS terminator), cauliflower mosaic virus CaMV 35S terminator, tml terminator, pea rbcS E9 terminator and nopaline and octopine synthase terminator (see, e.g., odell et al (I) ⁹⁸⁵ ) Nature 313:810; rosenberg et al (1987) Gene,56:125; guerineau et al (1991) mol. Gen. Genet,262:141; proudroot (1991) Cell,64:671; sanfacon et al Genes Dev.,5:141; mogen et al (1990) Plant Cell,2:1261; munroe et al (1990) Gene,91:151; ballad et al (1989) Nucleic Acids Res.17:7891; joshi et al (1987) Nucleic Acid Res., 15:9627).

Recombinant vectors containing the ZmYAB15 encoding gene expression cassette can be constructed using existing expression vectors.

In the above applications, the vector may be a plasmid, cosmid, phage or viral vector. The vector may specifically be a pBCXUN vector.

In the above application, the microorganism may be yeast, bacteria, algae or fungi. Wherein the bacterium may be an agrobacterium, such as agrobacterium EHA105.

In the above applications, none of the transgenic plant cell lines, transgenic plant tissues and transgenic plant organs include propagation material.

In the above application, the plant may be M1) or M2) or M3):

m1) monocotyledonous or dicotyledonous plants;

m2) a gramineous plant;

m3) corn.

In the above, the regulating drought resistance of the plant may be reducing drought resistance of the plant.

The application also provides any one of the following methods:

x1) a method of growing a drought resistance reduced plant comprising expressing ZmYAB15 in a recipient plant, or increasing the amount of ZmYAB15 in a recipient plant, or increasing the activity of ZmYAB15 in a recipient plant, to obtain a plant of interest having reduced drought resistance compared to said recipient plant;

x2) a method for reducing drought resistance in a plant comprising expressing ZmYAB15 in a recipient plant, or increasing the amount of ZmYAB15 in the recipient plant, or increasing the activity of ZmYAB15 in the recipient plant, to obtain a plant of interest having reduced drought resistance compared to said recipient plant, to achieve reduced drought resistance in the plant.

The methods of X1) and X2) can be accomplished by introducing a gene encoding ZmYAB15 into the recipient plant and allowing the gene to be expressed.

The coding gene may be the nucleic acid molecule of B1).

In the above method, the coding gene may be modified as follows before being introduced into the recipient plant to achieve better expression effect:

1) Modification and optimization are carried out according to actual needs so as to enable the genes to be expressed efficiently; for example, the codon of the coding gene of the present application may be changed to conform to plant preferences while maintaining the amino acid sequence thereof according to the codon preferred by the recipient plant; during the optimization process, it is preferable to maintain a certain GC content in the optimized coding sequence to best achieve high level expression of the introduced gene in the plant, wherein the GC content may be 35%, more than 45%, more than 50% or more than about 60%;

2) Modifying the gene sequence adjacent to the initiation methionine to allow efficient initiation of translation; for example, modifications are made using sequences known to be effective in plants;

3) Ligating to promoters expressed by various plants to facilitate expression thereof in plants; the promoter may include constitutive, inducible, chronologically regulated, developmentally regulated, chemically regulated, tissue-preferred, and tissue-specific promoters; the choice of promoter will vary with the time and space of expression requirements and will also depend on the target species; for example, a tissue or organ specific expression promoter, depending on the desired time period of development of the receptor; although many promoters derived from dicots have been demonstrated to be functional in monocots and vice versa, it is desirable to select dicot promoters for expression in dicots and monocot promoters for expression in monocots;

4) The expression efficiency of the gene of the application can be improved by connecting with a proper transcription terminator; e.g., tml derived from CaMV, E9 derived from rbcS; any available terminator known to function in plants may be ligated to the gene of the present application;

5) Enhancer sequences such as intron sequences (e.g., derived from Adhl and bronzel) and viral leader sequences (e.g., derived from TMV, MCMV and AMV) are introduced.

The recombinant expression vector may be introduced into plant cells by conventional biotechnological methods using Ti plasmids, plant virus vectors, direct DNA transformation, microinjection, electroporation, etc. (Weissbach, 1998,Method for Plant Molecular Biology VIII,Academy Press,New York,pp.411-463;Geiserson and Corey,1998,Plant Molecular Biology (2 nd Edition)).

The plant of interest is understood to include not only the first generation plants in which ZmYAB15 or its coding gene has been altered, but also their progeny. For the plant of interest, the gene may be propagated in that species, or may be transferred into other varieties of the same species, including particularly commercial varieties, using conventional breeding techniques. The plants of interest include seeds, calli, whole plants and cells.

In the above method, the recipient plant may be M1) or M2) or M3):

m1) monocotyledonous or dicotyledonous plants;

m2) a gramineous plant;

m3) corn.

ZmYAB15 is also within the scope of the present application.

The biological material also belongs to the protection scope of the application.

After the ZmYAB15 coding gene is overexpressed in corn, transgenic plants are easier to wilt under drought treatment conditions than control plants, and the survival rate is reduced after rehydration, so that the gene can reduce the drought resistance of plants, and the gene can be used for cultivating plants with reduced drought resistance so as to screen substances capable of improving the drought resistance of the plants.

Drawings

FIG. 1 is a phenotype of plants after drought treatment.

Figure 2 shows rehydration survival after drought treatment.

Detailed Description

The following detailed description of the application is provided in connection with the accompanying drawings that are presented to illustrate the application and not to limit the scope thereof. The examples provided below are intended as guidelines for further modifications by one of ordinary skill in the art and are not to be construed as limiting the application in any way.

The experimental methods in the following examples, unless otherwise specified, are conventional methods, and are carried out according to techniques or conditions described in the literature in the field or according to the product specifications. Materials, reagents, instruments and the like used in the examples described below are commercially available unless otherwise specified. The quantitative tests in the following examples were all set up in triplicate and the results averaged. In the following examples, unless otherwise specified, the 1 st position of each nucleotide sequence in the sequence listing is the 5 'terminal nucleotide of the corresponding DNA/RNA, and the last position is the 3' terminal nucleotide of the corresponding DNA/RNA.

The pBCXUN vector is an expression vector obtained by replacing the HYG gene (hptII, hygromycin resistance gene) of the pCXUN vector (GenBank: FJ905215.1, 06-JUL-2009) with the Bar gene (encoding phosphinothricin acetyltransferase) (nucleotides 284-835 in MG719235.1, 02-OCT-2018) and keeping the other nucleotides of pCXUN unchanged.

Example 1, zmYAB15 can regulate drought resistance of maize

In this example, a protein derived from maize B73 is found to regulate drought resistance of maize, the protein is denoted as ZmYAB15, the amino acid sequence of the protein is shown as SEQ ID No.3 in a sequence table, in maize B73, the genome sequence of ZmYAB15 is shown as SEQ ID No.1 in the sequence table, and the CDS sequence of the protein is shown as SEQ ID No.2 in the sequence table.

In SEQ ID No.1, the reading frame of the ZmYAB15 transcript is the 601-4346 nucleotide from the 5' end. The transcript of ZmYAB15 consists of 7 exons, namely 1289-1414, 1570-1746, 1918-2110, 2797-2845, 3120-3195, 3513-3587 and 3819-3908 of SEQ ID No.1, and introns are arranged among the exons.

1. Construction of recombinant vectors

The ZmYAB15 coding gene shown in SEQ ID No.2 of the sequence list is inserted into a pBCXUN vector to obtain a recombinant vector pBCXUN-ZmYAB15, and sequencing verification is carried out. In the recombinant vector pBCXUN-ZmYAB15, the expression of the exogenous DNA molecule is driven by the Ubi promoter to obtain the ZmYAB15 protein.

2. Construction of transgenic maize

The pBCXUN-ZmYAB15 obtained in the step 1 is introduced into an agrobacterium EHA105 strain to obtain recombinant strain EHA105/pBCXUN-ZmYAB15. Inoculating recombinant strain EHA105/pBCXUN-ZmYAB15 single colony into 2-3mL liquid culture medium containing 100 μg/mL kanamycin and 50 μg/mL rifampicin, shake culturing overnight at 28deg.C, transferring large amount of liquid culture medium containing 100 μg/mL kanamycin and 50 μg/mL rifampicin for the next day, shake culturing, collecting thallus after transferring several times, and re-suspending to OD ₆₀₀ Between 0.8 and 1.0, a recombinant Agrobacterium suspension was obtained. And (3) infecting young embryos of corn B73 scraped under aseptic conditions by using the obtained recombinant agrobacterium suspension, and then obtaining seedlings by inducing callus and screening herbicide glufosinate, and identifying to obtain transgenic plants. And obtaining a T3 generation after the transgenic plants are subjected to selfing propagation, and carrying out subsequent experiments.

Identification of transgenic plants: PCR amplification of genomic DNA of plants is performed by using primer pairs consisting of UbiP-seq (corresponding to the Ubi promoter) and NosR-seq (corresponding to the Nos terminator), so that plants with specific amplification products can be obtained as transgenic plants, and plants with specific amplification products cannot be obtained as non-transgenic plants. The primer sequences used were as follows:

UbiP-seq：TTTTAGCCCTGCCTTCATACGC；

NosR-seq：AGACCGGCAACAGGATTCAATC。

wherein two different transgenic inbred lines are ZmYAB15-OE1 and ZmYAB15-OE2 respectively.

3. Drought resistance identification of transgenic corn

Corn to be tested: zmYAB15-OE1, zmYAB15-OE2, and maize B73 (WT).

The detection steps are as follows: 140g of soil is added into each small basin, water is added into a tray, 4T 3 generation seeds or corn B73 seeds are placed into each small basin, 50ml of soil is covered, the rest of water in the tray is poured out after the water is absorbed, about three days after seedling emergence, one seedling with the worst growth vigor is removed, 1L of water is added into the tray, the water is poured out after the water is absorbed, drought treatment is started, no watering is continued for 20 days, the drought treatment phenotype of the control (WT) and transgenic corn is observed, normal watering and cultivation are resumed for 7 days, and then the phenotype (figure 2) is observed and survival rate is counted. 7 pots of each maize were replicated.

FIG. 1 shows that transgenic maize grew less than control plants after drought treatment and leaf wilting was stronger than control plants, and FIG. 2 shows that transgenic maize survived poorly after drought treatment with survival rates of ZmYAB15-OE1 and ZmYAB15-OE2 of 0 and 9.5%, respectively, all significantly lower than control plants (42.9% survival), indicating that transgenic maize was more sensitive to drought than control plants.

The present application is described in detail above. It will be apparent to those skilled in the art that the present application can be practiced in a wide range of equivalent parameters, concentrations, and conditions without departing from the spirit and scope of the application and without undue experimentation. While the application has been described with respect to specific embodiments, it will be appreciated that the application may be further modified. In general, this application is intended to cover any variations, uses, or adaptations of the application following, in general, the principles of the application and including such departures from the present disclosure as come within known or customary practice within the art to which the application pertains. The application of some of the basic features may be done in accordance with the scope of the claims that follow.

Sequence listing

<110> Chinese university of agriculture

<120> protein for controlling drought resistance of corn and use thereof

<160> 3

<170> PatentIn version 3.5

<210> 1

<211> 4946

<212> DNA

<213> corn (Zea mays L.)

<400> 1

agggtaggaa gtacgtacat atatacctgg tctctcggcc ccatttaagc cttggttcca 60

ccgtacccat gccgaggaaa atcgagccag accacggtca ggcttttgtc ttgcgaaaac 120

gcgaagaaaa gaagcacgcc gcacgcgtac cagagccctt ctcgcgccgg accggtcaga 180

tcggccgtgc tggcctgcgc ggctacacga cgcgcgcgtc caggcacatt gatgagaagt 240

atatatacaa gtacgtacgt acgtacaaag ggaacgtgcg cgcgtgcgta catgacaccc 300

acgtactatg gtgcctgcac ctcacctcac ctcacctctt gcgcgcggcc ggagagcagc 360

agagagggga gaccgcgcgc atcgatcgtg gcacaggttg ggggggccgg acgatcccca 420

ccactcgtcc cgcttttcca gtcggcaaca gtctcgagat gccgaccgcc ggtgcagaca 480

agctaggggc cggggggagg tagcgtgctg cctgctttgc ccccgtcaca tgtaccaacg 540

gctccacgct cgctttccct ttgtccaggg gccatatatc caccgcgctg tccgcggtcc 600

acacacccca tgccttgcct cctcctcctg cctgcctgcc tgcctttccc tcgccgctcg 660

cggcaagatc cacatgcagt agtaggagta ggagcagcaa gcaacaaggg agggagggag 720

cgaggtcttg ggctctctct tgttagttcc tgcgtgcttg ccctgccctg gatcggcttt 780

caaatcggga acaacgacag atccgaggct ttctcgaatc gggcaggtcc atcaaatcgc 840

aagtttagcc agccccagcc ccagccccag ccccaggcac agccagctag ccgtcaaatc 900

actcacactc atcaaatcaa aggggaccaa aaggatatct cctggctcct gcccctctcc 960

cctctcctag ctactcgtcg tataaaattc ccctttcgac tctcccccct ccccacacac 1020

acctgactct cctccatccc ctcctccgat ctctcaccac gctcctcttc tgctaccttg 1080

ctttccctgc tgcgatccgt tccttgccct attagtatta gctagccggc cggccggatc 1140

gatcgagcaa ggaggggcgc cccagcagtc ctcgtcggat cgatccatct cgtcgacagc 1200

gtcagcgcct caggtggatc ggagcggagg tggtggaccg gagctagtag ctgccagcca 1260

cacatttcgg tcgtcggaac cacatatcat gatgtcgtcc tcgtcctcgt cctcgtcgtc 1320

ggccgcctgc tgcttcccgc tggaccacct cgcgccgtcc cccaccgagc agctctgcta 1380

cgtgcactgc aactgctgcg acaccatcct cgccgtacgt gcatgcggcg ccccactctc 1440

acctataccg acactcgcac acaccagcta gcctctccca cttcttcttc tagctagcgc 1500

agcagccagc agcgcacgca gtgttgactg attcgtctct cttcgatctc tatatgcatg 1560

cgcgcgcagg tcggcgtgcc ttgcagcagc ctgttcaaga cggtgacggt gcgttgtggc 1620

cactgcgcca acctactctc cgtcaacctc cgcggcctcc tgctcccgcc cgccgcgccg 1680

gcgccgccca accatctcaa cttcgctcac tccttgctgt cacccacatc cccgcatggc 1740

ctcttggtat gtaactatgg accaaaaccc atccagctcg ttcgttcaac tagtttaaca 1800

tgcatgcact ttaacctgct agtgcatgtt tagtttcgtc gtccctagtt agcctcttca 1860

tataccactg agatcagtca gtgtgtgtgt gtgacttgct tgcttgcttg cgcgcaggac 1920

gagttggctc tccagcaggc gccgagcttc ctgatggagc aggcgagtgc caacctgagc 1980

agcaccatga cgggccgcag cagcaacagc agctgcgcca gcaacctgcc gccgccggcg 2040

ccgatgcctg cagctcagcc tgtgcagcag gaagccgaac tgcccaagac cgccccgtcg 2100

gtaaacaggc gtaaggactt cttgttcttt ctgtttccaa tctttggatc ccccatctct 2160

ctctctctct ctctctctct ctctctctct ctctctctct ctcttctgtt ttttattact 2220

ttctgcttta cttttccgaa ccaagggtat ttatcactgc actaatcaaa gacccaaagc 2280

cattatattg gcgtgatgcg gatggggatg aactcaggaa agtgtctgaa gaggatgagt 2340

aggggcaagc agatcaccgt ttgtcgtgct ggaacaataa caacaacttg atttgtgcat 2400

aaatagaact ggatgcatgg acgacgacac aatcccatcc tagcttatta tttcgtgcag 2460

tgttaggata aattacatat atatatgggt ttacaaggtt ataatagatt acatgagcat 2520

ttttcaccgc gtccagggaa catatatata tggatttgta taaaaaaagg gggaccttat 2580

cagggtcgat ggatggccta gctagcgaga catacaatca tgtcatgtta tagccagagg 2640

ttgcccatat ccatcgctct tcagttgcaa acagtcggca tgcatagtgt agtagttgct 2700

gctagctcta gatagataag cgtacccatt cgcccatgtt ccctacggat gcatgtatgc 2760

tgactcaaat ctaccctaca aaatctaact tcgcagctcc ggagaagcgg cagagagtcc 2820

cgtcggcata caaccggttc atcaagtaag attttgtgat aaaagtctct ctgtgcatgc 2880

attttccccc tttaaaaacc cttgtttgta tactctcata ttcacctcat ccatatgtat 2940

tgaagtggat tagagcgtaa attagtttaa gtttcagctc aagacatatg gattgagatg 3000

aactataaat acgagagtat ccaaacgaga tcagatagct ctagtatagt atgcgccact 3060

ctctgcatgc tcatcgtctt cttgctagta aatgtgtgct gctggctgcg tgcacacagg 3120

gacgagatcc agcgcatcaa ggccggcaac ccggacatca cccaccggga ggcgttcagc 3180

gcagccgcaa agaatgtaac tttctttcgt cttcttctcc agacgcatat atgcacagca 3240

gtgacaacta ctcgtggcta gcttgctgtc tgcgtgtctt gtgatttgac ccatcacttt 3300

acaatgtcag catggacaca gcgacaccta gtgtctggtg ctggctgctt gcgcctggtg 3360

tggtctgtgg ctagcttgct gccccaccgg cacagcgtcg tacagacgat acagctagcg 3420

ttcgttttca gtatttgctt ccaagtactc ggtcggtcgt acgtacccta ctaatgttgc 3480

attgcctttc tctgatttgt gttcataatc agtgggccca tttcccacac atccacttcg 3540

gcctcatgcc ggaccagggc ctcaagaaga ccttcaagac tcatcaggta tcagagctga 3600

cactgaaacc ctaataactc gatgtatatt ttgtatgcct tgaaatcctc acttgacact 3660

ttgacggagt tcaggtcatc cagcagcaac actatattac cacatgcttc ttttaagttt 3720

tttttttatt attcgaagtt ttgtagtact ttgtaagcac gcaccgtacc ccggccggcc 3780

tctaaaattt tggctggttc ctgggctgtg acatacagga tggcgctgaa gacatgctac 3840

tcaaagacga tctctacgcc gcagcggctg cagctgcagc agccaacatg ggcatcactc 3900

cattctaata actctggtta ggcatgtgtt ctgaatcagc tggcacgtcg tagtggctat 3960

cgccctttgt gctgcattct agggtttcgt ctcagatctt gatctcaaca tcgatcgtgt 4020

gtcgtttaat tagagtactt tccagtgtgg ggatagaaat ggagaataat caagttagag 4080

cgcgcatcaa gtaggccact gatgatccgt aggtacagac ttgattgtct cctctatata 4140

tcccttttca tttcatgttc tctctctttt tcctattttt tgttcctgga ccccttcaac 4200

tcgccagtag agagctcgtt gtgcctctcc aatcccatgt ttgcgtatgt ctgctcaaca 4260

acatgttttt accatggaga acaagttgaa catctgttag atctatcaag agtgtcatat 4320

atatgttcca tcgtccttta ccatgtactt ctactactac tatgtggttt ctttcaacat 4380

atagggttat attgcgttgt tgatttgcgg ttaatgtagt tgtgaaaact tgcatagtca 4440

ctattgcttg gacaattcaa aataatagtc tacatataaa taggcacaat atttccatat 4500

ggagatttta aaaataataa tgcaggtaat taattttagt tttacttgat tgatcgacga 4560

tacactgtcc ccttggaaat gaaactaaga tggaaaatca aattgaatca caaatggatc 4620

gactataatg tttaaaactt cttttggaaa tgtacaattt aaagattaac ttaggaataa 4680

caattatgtt tctctttgtc aatcaaaaag tgacctgcaa aataatagca tggttagatc 4740

tatgcactgt tcctttgatg ggaaatcaac attgaatgta aggattaatt aaatccatga 4800

attaactata ttgtttagtt cagtagtatg taaattaaat atagaagtgc gggagtgtac 4860

cccgcacaca catagacaca cgctcacaaa gtattttctc cacttgaaga acttaaattt 4920

gtttaaaaac atcatggtaa atggag 4946

<210> 2

<211> 786

<212> DNA

<213> corn (Zea mays L.)

<400> 2

atgatgtcgt cctcgtcctc gtcctcgtcg tcggccgcct gctgcttccc gctggaccac 60

ctcgcgccgt cccccaccga gcagctctgc tacgtgcact gcaactgctg cgacaccatc 120

ctcgccgtcg gcgtgccttg cagcagcctg ttcaagacgg tgacggtgcg ttgtggccac 180

tgcgccaacc tactctccgt caacctccgc ggcctcctgc tcccgcccgc cgcgccggcg 240

ccgcccaacc atctcaactt cgctcactcc ttgctgtcac ccacatcccc gcatggcctc 300

ttggacgagt tggctctcca gcaggcgccg agcttcctga tggagcaggc gagtgccaac 360

ctgagcagca ccatgacggg ccgcagcagc aacagcagct gcgccagcaa cctgccgccg 420

ccggcgccga tgcctgcagc tcagcctgtg cagcaggaag ccgaactgcc caagaccgcc 480

ccgtcggtaa acaggcctcc ggagaagcgg cagagagtcc cgtcggcata caaccggttc 540

atcaaggacg agatccagcg catcaaggcc ggcaacccgg acatcaccca ccgggaggcg 600

ttcagcgcag ccgcaaagaa ttgggcccat ttcccacaca tccacttcgg cctcatgccg 660

gaccagggcc tcaagaagac cttcaagact catcaggatg gcgctgaaga catgctactc 720

aaagacgatc tctacgccgc agcggctgca gctgcagcag ccaacatggg catcactcca 780

ttctaa 786

<210> 3

<211> 261

<212> PRT

<213> corn (Zea mays L.)

<400> 3

Met Met Ser Ser Ser Ser Ser Ser Ser Ser Ser Ala Ala Cys Cys Phe

1 5 10 15

Pro Leu Asp His Leu Ala Pro Ser Pro Thr Glu Gln Leu Cys Tyr Val

20 25 30

His Cys Asn Cys Cys Asp Thr Ile Leu Ala Val Gly Val Pro Cys Ser

35 40 45

Ser Leu Phe Lys Thr Val Thr Val Arg Cys Gly His Cys Ala Asn Leu

50 55 60

Leu Ser Val Asn Leu Arg Gly Leu Leu Leu Pro Pro Ala Ala Pro Ala

65 70 75 80

Pro Pro Asn His Leu Asn Phe Ala His Ser Leu Leu Ser Pro Thr Ser

85 90 95

Pro His Gly Leu Leu Asp Glu Leu Ala Leu Gln Gln Ala Pro Ser Phe

100 105 110

Leu Met Glu Gln Ala Ser Ala Asn Leu Ser Ser Thr Met Thr Gly Arg

115 120 125

Ser Ser Asn Ser Ser Cys Ala Ser Asn Leu Pro Pro Pro Ala Pro Met

130 135 140

Pro Ala Ala Gln Pro Val Gln Gln Glu Ala Glu Leu Pro Lys Thr Ala

145 150 155 160

Pro Ser Val Asn Arg Pro Pro Glu Lys Arg Gln Arg Val Pro Ser Ala

165 170 175

Tyr Asn Arg Phe Ile Lys Asp Glu Ile Gln Arg Ile Lys Ala Gly Asn

180 185 190

Pro Asp Ile Thr His Arg Glu Ala Phe Ser Ala Ala Ala Lys Asn Trp

195 200 205

Ala His Phe Pro His Ile His Phe Gly Leu Met Pro Asp Gln Gly Leu

210 215 220

Lys Lys Thr Phe Lys Thr His Gln Asp Gly Ala Glu Asp Met Leu Leu

225 230 235 240

Lys Asp Asp Leu Tyr Ala Ala Ala Ala Ala Ala Ala Ala Ala Asn Met

245 250 255

Gly Ile Thr Pro Phe

260

Claims (6)

1. Any of the following uses of a protein or a substance regulating the content of said protein:

d1 Reducing drought resistance of corn;

d2 Preparing a product for reducing drought resistance of corn;

the protein is A1), A2) or A3) as follows:

a1 A protein having an amino acid sequence of SEQ ID No. 3;

a2 A protein which is obtained by substituting and/or deleting and/or adding one or more amino acid residues for the amino acid sequence shown in SEQ ID No.3 in the sequence table and has the same function;

a3 A fusion protein obtained by ligating a tag to the N-terminal or/and the C-terminal of A1) or A2).

2. Use of a biological material related to the protein of claim 1 for any of the following:

d1 Reducing drought resistance of corn;

d2 Preparing a product for reducing drought resistance of corn;

the biomaterial is any one of the following B1) to B7):

b1 A nucleic acid molecule encoding the protein of claim 1;

b2 An expression cassette comprising the nucleic acid molecule of B1);

b3 A recombinant vector comprising the nucleic acid molecule of B1) or a recombinant vector comprising the expression cassette of B2);

b4 A recombinant microorganism comprising the nucleic acid molecule of B1), or a recombinant microorganism comprising the expression cassette of B2), or a recombinant microorganism comprising the recombinant vector of B3);

b5 A transgenic plant cell line comprising the nucleic acid molecule of B1) or a transgenic plant cell line comprising the expression cassette of B2);

b6 A transgenic plant tissue comprising the nucleic acid molecule of B1) or a transgenic plant tissue comprising the expression cassette of B2);

b7 A transgenic plant organ comprising the nucleic acid molecule of B1) or a transgenic plant organ comprising the expression cassette of B2).

3. The use according to claim 2, characterized in that: b1 The nucleic acid molecule is b 11) or b 12) or b 13) or b 14) as follows:

b11 A cDNA molecule or a DNA molecule of SEQ ID No.2 in the sequence table;

b12 A cDNA molecule or a DNA molecule shown in SEQ ID No.2 of the sequence Listing;

b13 A DNA molecule shown in SEQ ID No.1 of the sequence table;

b14 Under stringent conditions with a nucleotide sequence defined in b 11) or b 12) or b 13) and encoding a cDNA molecule or a genomic DNA molecule of a protein according to claim 1.

4. The method comprises the following steps:

x1) a method of breeding a maize with reduced drought resistance comprising expressing the protein of claim 1 in a recipient maize or increasing the amount of the protein of claim 1 in a recipient maize to obtain a maize of interest with reduced drought resistance compared to the recipient maize;

x2) a method for reducing drought resistance in maize comprising expressing the protein of claim 1 in a recipient maize or increasing the amount of the protein of claim 1 in the recipient maize to obtain a target maize having reduced drought resistance as compared to the recipient maize, whereby reduced drought resistance in maize is achieved.

5. The method according to claim 4, wherein: x1) and X2) is achieved by introducing into the recipient maize a gene encoding the protein of claim 1 and allowing expression of said gene.

6. The method according to claim 5, wherein: the coding gene is the nucleic acid molecule of B1) in claim 2 or 3.